Advanced Genetic Investigations of Unexplained Hyperinsulinemic Hypoglycemia in Children with Atypical Pancreatic Histology or NI-PHHS

Annette Rønholt Larsen*

*Corresponding author for this work

Research output: ThesisPh.D. thesis


Congenital hyperinsulinism (CHI) is a rare disease affecting around 1:28,000–50,000 individuals, characterized by abnormally high insulin secretion from the endocrine pancreas with resultant hyperinsulinemic hypoglycemia (HH). Diagnosis and treatment are essential to avoid brain damage or even death as a result of hypoglycemia. CHI is a heterogeneous disorder with differences in genetics, histology, clinical presentation, and treatment response. In non-syndromal congenital hyperinsulisme (CHI), pathogenic DNA mutations in at least 9 genes ABCC8, GCK, GLUD1, HADH, HK1, HNF1A, HNF4A, KCNJ11, and SLC16A1 have been described, most frequently in the KATP-channel genes ABCC8 and KCNJ11. Histologically, CHI has two major forms: 1) KATP-channel diffuse CHI form with beta cell hypertrophy with enlarged nuclei throughout the pancreas and 2) KATP-channel focal form with localized endocrine cell hyperplasia, leading to the formation of focal adenomatous hyperplasia. More rarely, histology is described with mutations in GCK and GLUD1. Atypical histology in non-syndromal CHI is defined in this thesis as non-KATP-channel diffuse CHI or non-KATP-channel focal CHI. The atypical histology and genetics in non-syndromal CHI have yet to be fully addressed and is the focus in five patients in this thesis. In later life, HH may be the result of insulinoma, insulinomatosis, or adultonset non-insulinoma persistent hyperinsulinemic hypoglycemia (NI-PHHS). In addition, a patient with histology corresponding to NI-PHHS is included in this thesis, however, without genetic explanation. We have applied a new sequencing method and laser-capture microdissection (LCM) to investigate for possible pathogenic variants in all the patients. Being able to determine genotype-histotype-phenotype correlations is vital to ensuring that families with HH can be advised and provided with the best possible treatment. The importance of identifying the KATP channel focal CHI disease is that these children can be cured with surgery.

In paper 1 we conducted a literature review to provide an overview of the research that has already been done, with an emphasis on the genotype-histotype-phenotype correlations. The review includes HH conditions with known histology, namely, KATP channel diffuse CHI, KATP channel focal CHI, GCK-CHI, GDH-CHI, BWS-CHI, Morphological mosaicism of pancreatic islets, insulinoma, insulinomatosis, and NI-PHHS.

In paper 2 we examined five histological atypical CHI patients. The patients underwent surgery with resection of 30–85% of the pancreas. The histological picture showed discrete changes incompatible with KATP channel diffuse CHI and KATP channel focal CHI. The study aimed to elucidate possibly pathogenic variants with new panelsequencing and LCM methods to determine the genotype-histotypephenotype correlation. Possible pathogenic variants with heterozygous germline de novo or low-grade mosaic HK1 intron 2 variants were found in three of the five patients. In Patient 3, one HK1 intron 2 variant was only found in the islets of Langerhans. In Patient 4, a paternal heterozygous likely pathogenic CACNA1D variant was found. We could not find any genetic explanation in Patient 5. We observed hints of a genotype-histotype-phenotype correlation in the patients, but further studies are required. The results from three other histological atypical patients from previous studies not mentioned in this thesis, will also be discussed in paper 2.

Paper 3 presented a patient with symptoms of HH from 9 years of age. Insulinoma was not detected. The patient underwent partial pancreatic resections at the age of 12 and again at the age of 14 years, which led to remission of the disorder. The study aimed to examine the genetics with a new panel sequencing and LCM. The histology showed enlarged islets of Langerhans, consistent with NI-PHHS. We could not find any possible pathogenic variants in either whole-pancreatic tissue or in isolated islets of Langerhans. The genotype could not be established despite multiple investigations.

Paper 4 draws attention to an alignment error caused by sequencing on two different sequencing platforms NextSeq 500/550 and NovaSeq 6000, respectively. When we sequenced the DNA from the pancreatic tissue from our patient from paper 3, we identified a somatic deletion and a point mutation in RNF40. However, we had to sequence the DNA sample again due to the subsequent RNA sequencing and analysis, which showed no difference in RNA expression compared to the control sample. Performing DNA sequencing on a newer machine, NovaSeq 6000, we were not able to replicate the results from the NextSeq 500/550 sequencing. We think that the difference may have been caused by the paired-end reads length as the NovaSeq 6000 platform provides a better alignment to the reference genome due to the longer base lengths. Researchers must be informed about the discrepancy between the two platforms to avoid possible misdiagnosis.
Original languageEnglish
Awarding Institution
  • University of Southern Denmark
  • Christesen, Henrik B Thybo, Principal supervisor
  • Detlefsen, Sönke, Co-supervisor
  • Brusgaard, Klaus, Co-supervisor
  • Andersen, Ditte Caroline, Co-supervisor
Date of defence3. May 2024
Publication statusPublished - 14. Mar 2024

Note re. dissertation

Print copy of the thesis is restricted to reference use in the Library.


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